Allogeneic hematopoietic stem cell transplantation (allo-HSCT) is the only curative treatment for patients with relapsed/refractory leukemia. The therapeutic benefits of allo-HSCT for hematologic malignancies are primarily derived from a graft-versus-leukemia (GvL) effect that is mediated by mature donor T cells present in the bone marrow graft. Unfortunately, the same donor T cells that mediate the beneficial GvL effect can also cause graft-versus-host disease (GvHD), the major life-threatening complication of allo-HSCT. Managing the threat of GvHD while maximizing the beneficial GvL effect would broaden the scope and usefulness of allo-HSCT procedures, CD4+CD25+FOXP3+ regulatory t cells (Tregs) have been shown to prevent GvHD in preclinical studies by suppressing alloreactive donor T cells without sacrificing GvL, thereby providing a promising treatment option. Unfortunately, several limitations have prevented the routine clinical use of Tregs: 1) the low circulating numbers of Tregs in peripheral blood, 2) loss of suppressor activity following in vitro expansion and 3) the lack of Treg-specific surface markers necessary to purify in vitro expanded Tregs. We previously reported that the DNA methyltransferase inhibitor azacitidine (AzaC)-induced Foxp3 expression and increased donor Tregs in vivo, thereby mitigating GvHD without abrogating GvL in a murine allo-HSCT model. Surprisingly, we found that AzaC-mediated suppression of GvHD was independent of Foxp3, the master regulator of Treg function. We identified three candidate genes that are highly upregulated by AzaC in anti-CD3/CD28 bead- and APC-activated CD4+/CD25- T cells and which might be responsible for the suppressor function of AzaC-induced Tregs based on genome-wide RNA profiling analyses. We hypothesize that AzaC will induce similar immunomodulatory effects in human T cells. In this SPORE project we will assess the safety and efficacy of AzaC in patients with AML and MDS undergoing allogeneic stem cell transplant (Aim 1) and will perform mechanistic studies to determine how AzaC and other DNMT1 inhibitors exert their immunomodulatory effects on GvHD and GvL in vitro and in vivo. (Aim 2).